U.S. patent number 5,518,988 [Application Number 08/253,951] was granted by the patent office on 1996-05-21 for method of counteracting an ethylene response in plants.
This patent grant is currently assigned to North Carolina State University. Invention is credited to Sylvia M. Blankenship, Edward C. Sisler.
United States Patent |
5,518,988 |
Sisler , et al. |
May 21, 1996 |
Method of counteracting an ethylene response in plants
Abstract
A method of inhibiting an ethylene response in a plant is
disclosed herein. The method comprises applying to the plant an
effective ethylene response-inhibiting amount of cyclopropene,
1.1.1. propellane, or derivatives thereof. Also disclosed are
methods of inhibiting abscission in plants and methods of
prolonging the life of cut flowers.
Inventors: |
Sisler; Edward C. (Raleigh,
NC), Blankenship; Sylvia M. (Apex, NC) |
Assignee: |
North Carolina State University
(Raleigh, NC)
|
Family
ID: |
22962324 |
Appl.
No.: |
08/253,951 |
Filed: |
June 3, 1994 |
Current U.S.
Class: |
504/114; 504/115;
504/320; 504/326; 504/353; 504/356; 504/357 |
Current CPC
Class: |
A01N
3/02 (20130101); A01N 27/00 (20130101); A01N
33/04 (20130101); A01N 35/08 (20130101) |
Current International
Class: |
A01N
3/00 (20060101); A01N 3/02 (20060101); A01N
27/00 (20060101); A01N 43/48 (20060101); A01N
45/00 (20060101); A01N 45/02 (20060101); A01N
003/02 (); A01N 027/00 (); A01N 029/04 (); A01N
033/04 () |
Field of
Search: |
;504/114,115,320,326,353,356,357 |
References Cited
[Referenced By]
U.S. Patent Documents
Other References
M C. Pirrung; Proposal to the Fred C. Gioeckner Foundation (1991).
.
Pirrung et al. "Ethylene Biosynthesis, Aminocyclopropene carboxylic
acid", J. Chem. Soc., Chem. Commun., (13), 857-859, 1989. .
Wheeler et al., "Synthesis of 1-aminocyclopropene carboxylic acid",
J. Org. Chem., 52(22) 4875-4877, 1987..
|
Primary Examiner: Robinson; Allen J.
Assistant Examiner: Bembenick; Brian G.
Attorney, Agent or Firm: Bell, Seltzer, Park &
Gibson
Government Interests
This invention was made with government support under Grant No.
91-37304-65 awarded by the U.S. Department of Agriculture. The
government has certain rights in the invention.
Claims
That which is claimed is:
1. A method of inhibiting an ethylene response in a plant,
comprising applying to the plant an effective ethylene
response-inhibiting amount of a compound of Formula I: ##STR3##
wherein: n is the number 1; and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy,
amino and carboxy.
2. A method according to claim 1, wherein n is 1.
3. A method according to claim 1, wherein said compound is
cyclopropene.
4. A method according to claim 1, wherein said applying step is
carried out by contacting said plant to a gas of said compound.
5. A method according to claim 1, wherein said applying step is
carried out by spraying said plant with a solution comprising said
compound.
6. A method according to claim 1, wherein said applying step is
carried out by contacting said plant to a solid comprising said
compound.
7. A method according to claim 1, wherein said ethylene response is
fruit ripening.
8. A method according to claim 7, wherein said fruit is selected
from the group consisting of tomatoes, apples, bananas, pears,
papaya, mangoes, peaches, apricots, nectarines, kiwi, pineapple,
persimmon, melons, pineapple, persimmon, berries, genus Cucumis,
green beans and avocados.
9. A method according to claim 1, wherein said ethylene response is
vegetable ripening.
10. A method according to claim 9, wherein said vegetable is
selected from the group consisting of lettuce, spinach, cabbage,
potatoes, carrots, onions, basil, oregano, dill, soybean, lima
beans, peas, corn, broccoli, cauliflower, and asparagus.
11. A method according to claim 1, wherein said ethylene response
is flower senescence.
12. A method according to claim 11, wherein said flower is selected
from the group consisting of azalea, hydrangea, hibiscus,
snapdragons, poinsettias, cactus, begonias, roses, tulips,
daffodils, petunias, carnations, lily, gladiolias, alstromeria,
anemone, columbine, avalia, aster, bouganvillea, camellia,
bellflower, cockscomb, false cypress, chrysanthemum, clematis,
cyclamen, freesia, and orchids.
13. A method of inhibiting abscission in a plant, comprising
applying to the plant an effective abscission-inhibiting amount of
a compound of Formula I: ##STR4## wherein: n is the number 1;
and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy,
amino and carboxy.
14. A method according to claim 13, wherein n is 1.
15. A method according to claim 13, wherein said compound is
1-methylcyclopropene.
16. A method according to claim 13, wherein said compound is
cyclopropene.
17. A method according to claim 13, wherein said applying step is
carried out by contacting said plant to a gas of said compound.
18. A method according to claim 13, wherein said applying step is
carried out by spraying said plant with a solution comprising said
compound.
19. A method according to claim 13, wherein said applying step is
carried out by contacting said plant to a solid comprising said
compound.
20. A method according to claim 13, wherein the plant is selected
from the group consisting of cotton, apple, pear, cherry, pecan,
grape, olive, coffee, snapbean, weeping fig, dormant seedlings,
privet, photinea, holly, ferns, schefflera, aglaonema, cotoneaster,
bar berry, waxmyrtle, abelia, acacia, and bromeliades.
21. A method of prolonging the life of a cut flower, comprising
applying to the cut flower an effective life-prolonging amount of a
compound of Formula I: ##STR5## wherein: n is a number from 1 to 2;
and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy,
amino and carboxy.
22. A method according to claim 21, wherein n is 1.
23. A method according to claim 21, wherein said compound is
1-methylcyclopropene.
24. A method according to claim 21, wherein said compound is
cyclopropene.
25. A method according to claim 21, wherein said compound is
3,3-dimethylcyclopropene.
26. A method according to claim 21, wherein said applying step is
carried out by contacting said plant to a gas of said compound.
27. A method according to claim 21, wherein said applying step is
carried out by spraying said plant with a solution comprising said
compound.
28. A method according to claim 21, wherein said applying step is
carried out by contacting said plant to a solid comprising said
compound.
29. A method of inhibiting an ethylene response in a plant,
comprising applying to the plant an effective ethylene
response-inhibiting amount of a compound of Formula I: ##STR6##
wherein: n is a number from 1 to 2; and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C1 to C4 alkyl, hydroxy, halogen, alkoxy, and
carboxy.
30. A method according to claim 29, wherein said compound is
3,3-dimethylcyclopropene.
31. A method of inhibiting an ethylene response in a plant,
comprising applying to the plant an effective ethylene
response-inhibiting amount of a compound of Formula I: ##STR7##
wherein: n is a number from 1 to 2; and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy, and
amino.
32. A method of inhibiting abscission in a plant, comprising
applying to the plant an effective abscission-inhibiting amount of
a compound of Formula I: ##STR8## wherein: n is a number from 1 to
2; and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy, and
carboxy.
33. A method according to claim 32, wherein said compound is
3,3-dimethylcyclopropene.
34. A method of inhibiting abscission in a plant, comprising
applying to the plant an effective abscission-inhibiting amount of
a compound of Formula I: ##STR9## wherein: n is a number from 1 to
2; and
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy, and
amino.
35. A method of inhibiting an ethylene response in a plant,
comprising applying to the plant an effective ethylene
response-inhibiting amount of 1-methylcyclopropene.
36. A method according to claim 35, wherein said applying step is
carried out by contacting said plant to a gas of said compound.
37. A method according to claim 35, wherein said applying step is
carried out by spraying said plant with a solution comprising said
compound.
38. A method according to claim 35, wherein said applying step is
carried out by contacting said plant to a solid comprising said
compound.
39. A method according to claim 35, wherein said ethylene response
is fruit ripening.
40. A method according to claim 39 wherein said fruit is selected
from the group consisting of tomatoes, apples, bananas, pears,
papaya, mangoes, peaches, apricots, nectarines, kiwi, pineapple,
persimmon, melons, pineapple, berries, genus Cucumis, green beans
and avocados.
41. A method according to claim 35, wherein said ethylene response
is vegetable ripening.
42. A method according to claim 41, wherein said vegetable is
selected from the group consisting of lettuce, spinach, cabbage,
potatoes, carrots, onions, basil, oregano, dill, soybean, lima
beans, peas, corn, broccoli, cauliflower, and asparagus.
43. A method according to claim 35, wherein said ethylene response
is flower senescence.
44. A method according to claim 43, wherein said flower is selected
from the group consisting of azalea, hydrangea, hibiscus,
snapdragons, poinsettias, cactus, begonias, roses, tulips,
daffodils, petunias, carnations, lily, gladiolias, alstromeria,
anemone, columbine, avalia, aster, bouganvillea, camellia,
bellflower, cockscomb, false cypress, chrysanthemum, clematis,
cyclamen, freesia, and orchids.
Description
FIELD OF THE INVENTION
The present invention generally relates to plant growth regulation,
and particularly relates to methods of inhibiting various ethylene
responses by applying cyclopropene, 1.1.1. propellane, or
derivatives thereof to the plant.
BACKGROUND OF THE INVENTION
Ethylene is known to mediate a variety of growth phenomena in
plants. See generally Fritz et al. U.S. Pat. No. 3,879,188. This
activity is understood to be achieved through a specific ethylene
receptor in plants. Many compounds other than ethylene interact
with this receptor: some mimic the action of ethylene; others
prevent ethylene from binding and thereby counteract its
action.
Many compounds which block the action of ethylene diffuse from the
binding site over a period of several hours. See E. Sisler and C.
Wood, Plant Growth Reg. 7, 181-191 (1988). These compounds may be
used to counteract ethylene action. A problem with such compounds,
however, is that exposure must be continuous if the effect is to
last for more than a few hours.
Photoaffinity labeling has been used in biological studies to label
binding sites in a permanent manner: usually by generating a
carbene or nitrene intermediate. Such intermediates are very
reactive and react rapidly and indiscriminately with many things. A
compound already bound, however, would react mostly to the binding
site. In a preliminary study, it was shown that cyclopentadiene was
an effective blocking agent for ethylene binding. See E. Sisler et
al., Plant Growth Reg. 9, 157-164 (1990). Methods of combatting the
ethylene response in plants with diazocyclopentadiene and
derivatives thereof are disclosed in U.S. Pat. No. 5,100,462 to
Sisler and Blankenship.
SUMMARY OF THE INVENTION
The foregoing and other objects and aspects of the present
invention are explained in detail in the specification set forth
below.
A method of inhibiting an ethylene response in a plant is disclosed
herein. The method comprises applying to the plant an effective
ethylene response-inhibiting amount of cyclopropene or a derivative
thereof.
Another aspect of the present invention is a method of blocking
ethylene receptors in plants by applying cyclopropene or a
derivative thereof to the plants in an effective receptor-blocking
amount.
Also disclosed is a method of inhibiting abscission in a plant,
comprising applying to the plant an effective abscission-inhibiting
amount of cyclopropene or a derivative thereof.
Also disclosed is a method of prolonging the life of a cut flower,
comprising applying to the cut flower an effective life-prolonging
amount of cyclopropene or a derivative thereof.
The methods described herein may be carried out in any suitable
manner, such as by contacting the plant to cyclopropene gas or a
gas of a cyclopropene derivative, or by spraying the plant with a
solution comprised of cyclopropene or a derivative thereof. These
and other suitable methods of application are discussed in detail
below.
A second method of inhibiting an ethylene response in a plant
disclosed herein comprises applying to the plant an effective
ethylene response-inhibiting amount of 1.1.1. propellane or a
derivative thereof.
Another aspect of the present invention is a method of blocking
ethylene receptors in plants by applying 1.1.1. propellane or a
derivative thereof to the plants in an effective receptor-blocking
amount.
Also disclosed is a method of inhibiting abscission in a plant,
comprising applying to the plant an effective abscission-inhibiting
amount of 1.1.1. propellane or a derivative thereof.
Also disclosed is a method of prolonging the life of a cut flower,
comprising applying to the cut flower an effective life-prolonging
amount of 1.1.1. propellane or a derivative thereof.
Yet another method of inhibiting an ethylene response in a plant is
disclosed. The method includes applying to the plant an effective
ethylene response-inhibiting amount of a compound isolated from
light treated diazocyclopentadiene by Gas chromatography, wherein
the compound elutes at about 0.7 minutes from a 23% SP-1700 on
80/100 Chromosorb P AW.TM. glass column.
The present invention also contemplates methods of blocking
ethylene receptors in plants, inhibiting abscission in plants, and
prolonging the life of cut flowers which include applying to the
plant an effective ethylene response-inhibiting amount of a
compound isolated from diazocyclopentadiene by Gas chromatography,
wherein the compound elutes at about 0.7 seconds from a 23% SP-1700
on 80/100 Chromosorb P AW.TM. glass column.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a gas-chromatograph, the peak at 0.70 illustrating an
ethylene response-inhibiting compound isolated from
diazocyclopentadiene.
FIG. 2 shows a graph depicting the concentration of
1-methylcyclopropene needed to achieve protection against exogenous
ethylene as a function of treatment time.
FIG. 3 compares the measurement of ethylene production in flowers
treated with ethylene, with 1-methylcyclopropene, and untreated
flowers.
FIG. 4 shows the irreversible binding of the ethylene inhibitor on
the ethylene receptor.
DETAILED DESCRIPTION OF THE INVENTION
As noted above, in addition to cyclopropene and 1.1.1. propellane,
various derivatives of cyclopropene and 1.1.1. propellane may also
be used to carry out the methods set forth herein.
Derivatives of cyclopropene which may be used to carry out the
present invention are defined by Formula (I) below: ##STR1##
wherein:
n is a number from 1 to 4. Preferably n is a number from 1 to 2,
and most preferably n is 1.
R is selected from the group consisting of hydrogen, saturated or
unsaturated C.sub.1 to C.sub.4 alkyl, hydroxy, halogen, alkoxy,
amino and carboxy.
Derivatives of 1.1.1. propellane which may be used to carry out the
present invention include those defined by Formula (II) below:
##STR2## wherein n is from 1 to 3, preferably 1, and R is as
defined above with reference to Formula (I).
R groups are preferably substituted on the compound of formula (II)
on those carbon atoms which are covalently bound to two other
carbon atoms, rather than those carbon atoms which are covalently
bound to three other carbon atoms (i.e., to carbon atoms at the
apexes of the structure shown).
The term .-+.alkyl" as used herein refers to linear or branched,
saturated or unsaturated alkyl. Examples include, but are not
limited to, methyl, ethyl, propyl, isopropyl, and butyl. Alkyl
groups of the present invention are preferably linear and
saturated.
The term "plant" is used in a generic sense herein, and encompasses
woody-stemmed plants such as trees and shrubs. Plants to be treated
by the methods described herein include whole plants and any
portions thereof, such as field crops, potted plants, cut flowers
(stems and flowers), and harvested fruits and vegetables.
Plants treated by the methods of the present invention are
preferably treated with a non-phytotoxic amount of the active
compound.
The present invention can be employed to combat numerous different
ethylene responses. Ethylene responses may be initiated by either
exogenous or endogenous sources of ethylene. Ethylene responses
include, for example, the ripening and/or senescence of flowers,
fruits and vegetables, abscission of foliage, flowers and fruit,
the prolongation of the life of ornamentals such as potted plants,
cut flowers, shrubbery, and dormant seedlings, in some plants
(e.g., pea) the inhibition of growth, and in other plants (e.g.,
rice) the stimulation of growth.
Vegetables which may be treated by the method of the present
invention to inhibit ripening and/or senescence include leafy green
vegetables such as lettuce (e.g., Lactuea sativa), spinach (Spinaca
oleracea), and cabbage (Brassica oleracea), various roots, such as
potatoes (Solanum tuberosum) and carrots (Daucus), bulbs, such as
onions (Allium sp.), herbs, such as basil (Ocimum basilicum),
oregano (Origanum vulgare), dill (Anethum graveolens), as well as
soybean (Glycine max), lima beans (Phaseolus limensis), peas
(Lathyrus spp.), corn (Zea mays), broccoli (Brassica oleracea
italica), cauliflower (Brassica oleracea botrytis), and asparagus
(Asparagus officinalis).
Fruits which may be treated by the method of the present invention
to inhibit ripening include tomatoes (Lycopersicon esculentum),
apples (Malus domestica), bananas (Musa sapientum), pears (Pyrus
communis), papaya (Carica papaya), mangoes (Mangifera indica),
peaches (Prunus persica), apricots (Prunus armeniaca), nectarines
(Prunus persica nectarina), oranges (Citrus sp.), lemons (Citrus
limonia), lines (Citrus aurantifolia), grapefruit (Citrus
paradisi), tangerines (Citrus nobilis deliciosa), kiwi (Actinidia
chinenus), melons such as cantaloupe (C. cantalupensis) and musk
melon (C. melo), pineapple (Aranas comosus), persimmon (Diospyros
sp.), various small fruits including berries such as strawberries
(Fragaria), blueberries (Vaccinium sp.) and raspberries (e.g.,
Rubus ursinus), green beans (Phaseolus vulgaris), members of the
genus Cucumis such as cucumber (C. sativus), and avocados (Persea
americana).
Ornamental plants which may be treated by the method of the present
invention to inhibit senescence and/or to prolong flower life and
appearance (e.g., delay wilting), include potted ornamentals, and
cut flowers. Potted ornamentals and cut flowers which may be
treated with the present invention include azalea (Rhododendron
spp.), hydrangea (Macrophylla hydrangea) hybiscus (Hibiscus
rosasanensis), snapdragons (Antirrhinum sp.), poinsettia (Euphorbia
pulcherima), cactus (e.g. Cactaceae schlumbergera truncata),
begonias (Begonia sp.), roses (Rosa spp.), tulips (Tulipa sp.),
daffodils (Narcissus spp.), petunias (Petunia hybrida), carnation
(Dianthus caryophyllus), lily (e.g., Lilium sp.), gladiolus
(Gladiolus sp.), alstroemeria (Alstoemeria brasiliensis), anemone
(e.g., Anemone blanda), columbine (Aquilegia sp.), aralia (e.g.,
Aralia chinensis), aster (e.g., Aster carolinianus), bougainvillea
(Bougainvillea sp.), camellia (Camellia sp.), bellflower (Campanula
sp.), cockscomb (celosia sp.), falsecypress (Chamaecyparis sp.),
chrysanthemum (Chrysanthemum sp.), clematis (Clematis sp.),
cyclamen (Cyclamen sp.), freesia (e.g., Freesia refracta), and
orchids of the family Orchidaceae.
Plants which may be treated by the method of the present invention
to inhibit abscission of foliage, flowers and fruit include cotton
(Gossypium spp.), apples, pears, cherries (Prunus avium), pecans
(Carva illinoensis), grapes (Vitis vinifera), olives (e.g. Vitis
vinifera and Olea europaea), coffee (Coffea arabica), snapbeans
(Phaseolus vulgaris), and weeping fig (ficus benjamina), as well as
dormant seedlings such as various fruit trees including apple,
ornamental plants, shrubbery, and tree seedlings. In addition,
shrubbery which may be treated according to the present invention
to inhibit abscission of foliage include privet (Ligustrum sp.),
photinea (Photinia sp.), holly (Ilex sp.) ferns of the family
Polypodiaceae, schefflera (Schefflera sp.), aglaonema (Aglaonema
sp.), cotoneaster (Cotoneaster sp.), barberry (Berberis sp.),
waxmyrtle (Myrica sp.) abelia (Abelia sp.), acacia (Acacia sp.) and
bromeliades of the family Bromeliaceae.
Additional ethylene responses include those listed in Fritz et al.
U.S. Pat No. 3,879,188 at Column 3 line 62 through Column 6 line
65, the disclosure of which is incorporated herein by reference in
its entirety.
The active compound of the present invention can be applied to
plants by any suitable means. They may be applied alone, or in
combination with inert carriers. The active compound may be applied
alone in gaseous, liquid, or solid form, by contacting the compound
to the plant to be treated. Additionally the active compound may be
converted to the salt form, and then applied to the plants.
Alternatively, the compound may be applied with a inert carrier.
Suitable solid carriers include dust. The active compound may also
be suspended in a liquid solution, as an organic solvent or an
aqueous solution. Similarly, the gaseous form of the compound may
be dispersed in an inert gaseous carrier to provide a gaseous
solution.
Numerous organic solvents may be used as a carrier for the active
compounds of the present invention, e.g., hydrocarbons such as
hexane, benzene, toluene, xylene, kerosene, diesel oil, fuel oil
and petroleum naphtha, ketones such as acetone, methyl ethyl ketone
and cyclohexanone, chlorinated hydrocarbons such as carbon
tetrachloride, esters such as ethyl acetate, amyl acetate and butyl
acetate, ethers, e.g., ethylene glycol monomethyl ether and
diethylene glycol monomethyl ether, alcohols, e.g., ethanol,
methanol, isopropanol, amyl alcohol, ethylene glycol, propylene
glycol, butyl carbitol acetate and glycerine.
Mixtures of water and organic solvents, either as solutions or
emulsions, can be also employed as carriers for the active
compound.
The active compounds can be applied as aerosols, e.g., by
dispersing them in air by means of a compressed gas such as
dichlorodifluoromethane or trichlorofluoromethane and other Freons,
for example.
The active compounds of the present invention can also be applied
with adjuvants or carriers such as talc, pyrophyllite, synthetic
fine silica, attapulgus clay (attaclay), kieselguhr, chalk,
diatomaceous earth, lime, calcium carbonate, bentonite, fuller's
earth, cottonseed hulls, wheat flour, soybean flour pumice,
tripoli, wood flour, walnut shell flour, redwood flour and
lignin.
It may be desirable to incorporate a wetting agent in the
compositions of the present invention. Such wetting agents may be
employed in both the solid and liquid compositions. The wetting
agent can be anionic, cationic or nonionic in character.
Typical classes of wetting agents include alkyl sulfonate salts,
alkylaryl sulfonate salts, alkyl sulfate salts, alkylamide
sulfonate salts, alkylaryl polyether alcohols, fatty acid esters of
polyhydric alcohols and the alkylene oxide addition products of
such esters, and addition products of long chain mercaptans and
alkylene oxides. Typical examples of such wetting agents include
the sodium alkylbenzene sulfonates having 10 to 18 carbon atoms in
the akyl group, alkylphenol ethylene oxide condensation products,
e.g., p-isooctylphenol condensed with 10 ethylene oxide units,
soaps, e.g., sodium stearate and potassium oleate, sodium salt of
propylnaphthalene sulfonic acid (di-2-ethylhexyl), ester of sodium
sulfosuccinic acid, sodium lauryl sulfate, sodium stearate and
potassium oleate, sodium salt of the sulfonated monoglyceride of
coconut fatty acids, sorbitan, sesquioleate, lauryl trimethyl
ammonium chloride, octadecyl trimethyl ammonium chloride,
polyethylene glycol lauryl ether, polyethylene esters of fatty
acids and rosin acids, e.g., Ethofat 7 and 13, sodium
N-methyl-N-oleyltaurate, Turkey Red oil, sodium dibutylnaphthalene
sulfonate, sodium lignin sulfonate (Marasperse N), polyethylene
glycol stearate, sodium dodecylbenzene sulfonate, tertiary dodecyl
polyethylene glycol thioether (Nonionic 218), long chain ethylene
oxide-propylene oxide condensation products, e.g., Pluronic 61
(molecular weight 1,000), sorbitan sesquioleate, polyethylene
glycol ester of tall oil acids, sodium octyl phenoxyethoxyethyl
sulfate, polyoxyethylene (20) sorbitan monolaurate ("Tween 20")
tris (polyoxyethylene) sorbitan monostearate ("Tween 60"), and
sodium dihexyl sulfosuccinate.
The solid, liquid, and gaseous formulations can be prepared by any
of the conventional procedures. Thus, the active ingredient, in
finely divided form if a solid, may be tumbled together with finely
divided solid carrier. Alternatively, the active ingredient in
liquid form, including solutions, dispersions, emulsions and
suspensions thereof, may be admixed with the solid carrier in
finely divided form.
The present invention is explained in greater detail in the
following non-limiting Examples. In these examples, .mu.l means
microliters; ml means milliliters; l means liters, cm means
centimeters; and temperatures are given in degrees Centigrade.
EXAMPLE 1
Isolation of Active Compound from Diazocyclopentadiene
Diazocyclopentadiene was placed in a glass container and exposed to
light. A VARIAN.TM. Gas chromatograph equipped with a flame
ionization detector and a 23% SP-1700 on 80/100 Chromosorb P AW.TM.
glass column with nitrogen as a carrier gas was used to separate
the mixture. Diazocyclopentadiene was analyzed at ambient
temperature. A chromatogram was produced and the elution of each
peak was timed using a stopwatch. An exemplary chromatogram is
reproduced as FIG. 1.
Thereafter, the detector flame was extinguished and a piece of
TYGON.TM. tubing was placed securely over the flame tip. The
opposite end of the tubing was fitted with the hub of a syringe
needle. Meanwhile, carnations were placed in glass jars containing
water. Each peak was then permitted to elute into a separate jar.
The procedure was repeated 5 times. Approximately 1 ml of pure
oxygen was added to the jar to compensate for the reduction in the
oxygen level due to the nitrogen carrier gas. The carnations were
maintained overnight. The next day, the carnations were aired.
Thereafter, the jars were resealed and ethylene gas was injected
into the jars. The flowers were exposed to the ethylene atmosphere
overnight. The next day the carnations were again aired and removed
from the jars. Flowers were placed in water at room temperature
until symptoms of senescence developed. Flowers that dried, closed
up and showed typical ethylene injury symptoms were judged to be
unprotected by a compound. Flowers that retained their fresh
appearance were judged to be protected from the ethylene by the
compound. The peak which eluted at about 0.70 minutes consistently
protected the carnations from ethylene injury. The retention time
on the column indicated that the active compound most likely
contained 3 or 4 carbon atoms.
EXAMPLE 2
Measurement of Ethylene Binding
Triplicate samples of 3 g cut carnations' petals were placed in a
2.51 desiccator containing .sup.14 C-ethylene-mercuric perchlorate
complex (110 mCi/mM) in a 25 ml Erlenmeyer flask. Then, an excess
of unsaturated lithium chloride was added to the ethylene-mercuric
perchlorate complex to release the gaseous ethylene. The mixture
was stirred for 6 minutes. To determine the amount of binding, 3 ml
of unlabelled ethylene was added in one desiccator. After 2 hours
of ethylene exposure, the desiccators were opened and the samples
were aerated for 4 minutes. Each sample was then placed in a 250 ml
jar. 0.2 Ml of mercuric perchlorate on a 0.5 cm.sup.2 piece of
fiber-glass filter in a scintillation vial was then added. After 18
hours, the scintillation vials were removed, scintillation fluid
added and the radioactivity counted for each sample.
EXAMPLE 3
Effect of 1-Methylcyclopropene and Dimethylcyclopropene in
preserving in vivo carnations exposed to high levels of
ethylene
The treatment of carnations with ethylene hastens the process of
senescence, producing a petal in-rolling phenomena (Halevy and
Mayak, 1981). Carnations at stage II, with low ethylene production
and no visible signs of senescence (Woodson, 1987) were treated
with 1-methylcyclopropene at different concentration for 6 hours
before adding 10 or 1000 ml/l of ethylene for 1 hour.
After 4 days, the carnations treated with only 2.5 nl/l of
1-methylcyclopropene looked like the control that did not have the
ethylene treatment. The minimal concentration of
1-methylcyclopropene preventing the ethylene process was the same
when flowers were treated with 1 ml/l of ethylene.
These results on in vivo carnations suggest that
1-methylcyclopropene acts as a potent inhibitor of ethylene
response in the same way as STS, DACP or NBD. However, the
concentration of 1-methylcyclopropene to protect the flower against
ethylene action is much lower than these other chemical
products.
EXAMPLE 4
Treatment time of 1-methylcyclopropene
1-Methylcyclopropene was added for various times before the
application of exogenous ethylene. FIG. 2 shows that the
concentration of 1-Methylcyclopropene needed to get a protection
against the effect of exogenous ethylene was inversely related to
the treatment time. Treatment with about 250 to 300 nl/l of
1-methylcyclopropene for five minutes was enough to protect the
flowers. Treated for 24 hours, with 0.5 nl/l protected against 1
ml/l of ethylene.
EXAMPLE 5
Irreversible binding of 1-methylcyclopropene on in vivo
carnations
Flowers were treated with 5 nl/l of product during 6 hours, then,
stored for 10 days at room temperature. One ml/l of ethylene was
then added over 18 hours. Ethylene had no effect on the treated
carnations 10 days after the 1-methylcyclopropene treatment (data
not shown). The binding of 1-methylcyclopropene on carnations seems
to be irreversible.
EXAMPLE 6
Effect of 1-methylcyclopropene on in vivo carnations to stop the
senescent process due to exogenous and endogenous ethylene
Flowers in a pre-senescent stage were treated with 5 nl/l of
1-methylcyclopropene and the senescent process was observed. An
untreated control flower, began to exhibit petal in-rolling one day
later (data not shown). The 1-methylcyclopropene treated flower did
not show a senescent process 15 days later.
The carnations need to have an exogenous ethylene exposure at least
for 6 hours to have a visible sign of senescence. Treatment with 3
nl/l of 1-methylcyclopropene is enough to stop that process.
Carnations treated only with ethylene exhibited petal in-rolling,
whereas carnations treated with ethylene and then
1-methylcyclopropene exhibited no visible sign of the phenomenon
(data not shown). 1-methylcyclopropene seems to reduce and prevent
the autocatalytic production of ethylene.
The ability of 1-methylcyclopropene to stop the senescence process
was observed by adding ethylene over 6 hours. The carnations began
to exhibit the in-rolling phenomena. Thereafter
1-methylcyclopropene was added. The process of senescence was
stopped by the binding of 1-methylcyclopropene on the ethylene
receptor and giving a molecular response.
EXAMPLE 7
Effect of 1-methylcyclopropene on ethylene production
Ethylene production was followed from the beginning of the ethylene
climacteric production. The control exhibited a rise in ethylene
production 4 days after beginning the experiment. Flowers first
treated with ethylene, exhibited a rise in ethylene production 2
days earlier. 1- Methylcyclopropene was applied. Thereafter
ethylene production was measured. As shown in FIG. 3, the measure
of the ethylene production observed in the control was considerably
lower than that observed in flowers treated with
1-methylcyclopropene.
EXAMPLE 8
Effect of 1-methylcyclopropene as a function of the stage of cut
carnations
Cut carnations at stage I and stage III were treated with
1-methylcyclopropene. For young carnations, the concentration of
1-methylcyclopropene giving a protective effect was about 1.25 and
2.5 nl/l. For these, the protection against ethylene was total. For
old carnations, rates between 2.5 and 5 nl/l of
1-methylcyclopropene were sufficient to give a response, but total
protection was not achieved. Total protection of older carnations
was only achieved with 10 nl/l of 1-methylcyclopropene (data not
shown).
EXAMPLE 9
Irreversible binding of 1-methylcyclopropene on the ethylene
receptor
To determine if the 1-methylcyclopropene acted on the ethylene
receptor, flowers were treated with 5 nl/l of 1-methylcyclopropene
and stored for 4 days at room temperature at 4.degree. C. before
the ethylene binding experiment. Carnation petals preincubated with
1-methylcyclopropene and control petals which had not been
incubated with 1-methylcyclopropene were then incubated in the
presence of .sup.14 C-ethylene. One sample representing the control
was incubated with a saturating concentration of unlabelled
ethylene in the presence of .sup.14 C-ethylene. The difference in
labelling of the 1-methylcyclopropene treated sample versus the
non-treated sample and the control indicates the specific binding
of ethylene. In the two cases, the ethylene binding was totally
inhibited (see FIG. 4). Flowers treated with 1-methylcyclopropene
did not bind ethylene.
EXAMPLE 10
Diffusion of .sup.3 H 1-methylcyclopropene on carnation petals
In order to label the ethylene receptor, 1-methylcyclopropene was
labelled with tritium and the specific activity obtained was 60
mCi/mM. An eventual diffusion of the compound was studied on
carnations. The flowers were treated with 1-methylcyclopropene, and
with large amounts of ethylene. Control flowers were treated with
1-methylcyclopropene alone. The diffusion was followed for 7 days.
Only the flowers which did not have the ethylene treatment showed a
little diffusion. When the experiment was made at 4.degree. C., the
diffusion was nonexistent. These results suggest that .sup.3 H
1-methylcyclopropene was permanently bound to carnation
tissues.
EXAMPLE 11
Effect of 1-methylcyclopropene on banana ripening
Bananas were individually placed in a 3 l jar. 1-Methylcyclopropene
was injected into the jar at a determined concentration. The plant
material was then aerated and 1 ml/l of ethylene was added for 12
hours. Controls were held in jars without chemical treatment.
Chlorophyll measurement was made as previously described in Sisler
and Wood, 1988). The experiment was done 7 days after
1-methylcyclopropene treatment.
The Ki obtained for chlorophyll disappearance was 40 ul/l when
experiments were done in the dark and 1.2 ul/l when they were done
in the light. The quantification of the effect of
1-methylcyclopropene was done by chlorophyll measurement. The Ki
obtained for 1-methylcyclopropene was 0.4 nl/l. The ripening of
1-methylcyclopropene treated bananas was prevented for about 15
days and fruits turned brown after this period.
EXAMPLE 12
Effect of 1-methylcyclopropene tomato seed germination and tomato
ripening
Tomato seeds were washed with NaOCl (10%) and rinsed with water.
The seeds were placed on wetted filter paper with 10 mM sodium
phosphate buffer at pH 5.8. The germination was done in the dark.
When the seedlings were about 1 mm height, 30 seeds by sample were
placed on wet filter paper in a 0.5 l jar. 1-Methylcyclopropene was
added over 24 hours before adding 10 .mu.l/l ethylene for 5
days.
When 1-methylcyclopropene was applied for 12 hours before the
ethylene treatment, only 10 nl/l of compound was enough to preserve
tomatoes. On these fruits, 1-methylcyclopropene had a temporary
effect to prevent the ripening, which occurred about 7 to 10 days
after the 1-methylcyclopropene treatment.
The foregoing is illustrative of the present invention and is not
to be construed as limiting thereof. The invention is defined by
the following claims, with equivalents of the claims to be included
therein.
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